Lentil (Lens culinaris Medikus) may have a potential to enhance the productivity of agroecosystems in dry areas where water and nutrients are limited. This study quantified soil water, residual soil nitrogen (N), and crop yields in lentil-based systems in comparison with continuous cereal and conventional summerfallow systems. A 3-yr cropping sequence study was conducted for three cycles in Saskatchewan (50.28°N, 107.79°W) from 2007 to 2011. On average, soil retained 187, 196 and 337 mm of water in the 0–1.2 m depth at crop harvest in 2008, 2009 and 2010, respectively. Summerfallow contained the same amounts of water as the cropped treatments at the harvest in 2009 and 2010. However, in 2008, summerfallow contained more soil water than the cropped treatments. The effect of lentil cultivar on soil water conservation varied with years; the cultivars Glamis, Laird and Sedley conserved highest amounts of soil water by the planting time of 2009 and 2010, but no differences were found among cultivars in 2011. Soil available N (NO3− + NH4+) at spring planting time was 50.4 kg ha−1 in the preceding lentil treatments, which was 44% higher compared with preceding barley or flax, but was 25% lower compared with preceding summerfallow. Lentil cultivars had a similar amount of soil residual N. Grain production in the 3-yr rotation averaged 6.3 t ha−1 per rotation for the wheat–lentil–durum system and 6.8 t ha−1 for the wheat–cereal–durum monoculture, averaging 36% greater compared with wheat–summerfallow–durum system. The lentil system increased total grain production through the access of residual soil water and biologically fixed N, whereas continuous cereal system relies on inorganic fertilizer input for yield. Summerfallow system relies on ‘mining’ the soil for nutrients. We conclude that the adoption of lentil systems will enhance grain production through the use of residual soil water and available N.

N2 fixation, measured as acetylene reduction, was studied in laboratory cultures and in natural assemblages (both as a mixed population and as individually picked colonies) of the heterocystous cyanobacteria Aphanizomenon sp. and Nodularia spp. from the Baltic Sea. During a diurnal cycle of alternating light and darkness, these organisms reduced acetylene predominantly during the period of illumination, although considerable activity was also observed during the dark period. In both laboratory cultures and natural populations N2 fixation was saturated below a photon flux density of 600 μm−2 s−1. In cyanobacterial blooms in the Baltic Sea, nitrogenase activity was mostly confined to the surface layers. Samples collected from greater depths did not possess the same capacity for acetylene reduction as samples from the surface itself, even when incubated at the photon flux density prevailing in surface waters. This suggests that, with respect to N2 fixation, Baltic cyanobacteria are adapted to the intensity of illumination that they are currently experiencing.

Availability of nitrogen (N) and phosphorus (P) might significantly affect N2 fixation in legumes. The interaction of N and P was studied in common bean (Phaseolus vulgaris), considering their effects on nodulation and N2 fixation, nitrate reductase activity, and the composition of N compounds in xylem sap. The effect of N on the uptake of P by plants was estimated by analysing rhizospheric pH and P concentration in xylem sap and in plant shoots. Inoculated bean plants were grown in pots containing perlite/vermiculite in two experiments with different amounts of P and N. In a third experiment, bean plants were grown on two soil types or on river sand supplied with different concentrations of N. At harvest, shoot growth, number of nodules and mass, and nitrogenase activity were determined. Xylem sap was collected for the determination of ureides, amino acids, nitrate and phosphate concentration. At low nitrate concentration (1 mM), increasing amounts of P promoted both nodule formation and N2 fixation, measured as ureide content in the xylem sap. However, at high nitrate concentration (10 mM), nodulation and N2 fixation did not improve with increased P supply. Glutamine and aspartate were the main organic N compounds transported in the xylem sap of plants grown in low nitrate, whereas asparagine was the dominant N compound in xylem sap from plants grown in high nitrate. Nitrate reductase activity in roots was higher than in shoots of plants grown with low P and high N. In both soils and in the sand experiment, increased application of N decreased nodule mass and number, nitrogenase activity and xylem ureides but increased the concentration of asparagine in xylem sap. Increasing P nutrition improved symbiotic N2 fixation in bean only at low N concentrations. It did not alleviate the inhibitory effect of high nitrate concentration on N2 fixation. A decrease in plant P uptake was observed, as indicated by a lower concentration of P in the xylem sap and shoots, correlating with the amount of N supplied. Simultaneously with the specific inhibition of N2 fixation, high nitrate concentrations might decrease P availability, thus inhibiting even further the symbiotic association because of the high P requirement for nodulation and N2 fixation.

Interactive effects of elevated atmospheric CO2 and soil N availability on N2 fixation and biomass production were examined using Gliricidia sepium, a tropical leguminous tree species. Our objective was to determine if elevated CO2 alters the inhibitory effects of soil N on N2 fixation, and whether the response of Gliricidia to elevated CO2 was a function of N source originating from either substrate N fertilizer or N2 fixation. We hypothesized that CO2 enrichment would ameliorate the inhibitory effects of N fertilization on seedling nodulation and N2 fixation through increased C partitioning to nodules. Seedlings were grown from seed for 100 d in growth chambers at either 350 or 700 μmol mol−1 CO2. Seedlings were inoculated with Rhizobium spp. and grown either with 0, 1 or 10 mM N fertilizer. The δ15N isotope-dilution technique was used to determine N source partitioning between N2 fixation and inorganic N fertilizer uptake. The addition of 10 mM N fertilizer significantly reduced nodule number and mass, specific nitrogenase activity, the specific rate of N2 fixation, and the proportion of plant N derived from N2 fixation. Elevated CO2, however, strongly ameliorated the inhibitory effects of N fertilization, indicating that increased C availability for nodule activity may partially offset the inhibition of N2 fixation caused by substrate N, as nodule sugar concentrations were stimulated with CO2 enrichment. This study clearly shows that elevated CO2 enhanced plant productivity and net N content of Gliricidia tree seedlings by stimulating N2 fixation. In addition, seedling biomass production was greatly enhanced by elevated CO2, regardless of whether plant N was derived from the substrate or from the atmosphere. We conclude from this study that CO2 enrichment mitigates the inhibitory effects of substrate N on nodule initiation and development and specific N2 fixation, either through increased C allocation to nodule production and activity, or through increased N demand by the plant for biomass production. This experiment with Gliricidia provides evidence for a positive feedback between increased atmospheric CO2 concentrations, C allocation to symbiotic N2-fixing bacteria, and plant C and N accumulation that may occur when N2-fixing plants are grown under conditions where substrate N may typically inhibit N2 fixation.

The nodulation status and nodule morphology of 62 taxa of Leguminosae in a rain forest in French Guiana are reported according to the taxonomy of the family. The N2-fixing species are then fitted into ‘functional groups’ according to their behaviour towards illumination, in order to evaluate their importance in the global dynamics of the stand. The results showed that 67% of the observed species were nodulated (50, 71 and 77% of the Caesalpiniaceae, Mimosaceae and Papilionaceae, respectively). In the Caesalpiniaceae, nodule-like structures were reported in the genus Crudia and in the species Senna quinquangulata, although this needs to be confirmed. All the nodules studied in this subfamily were astragaloid and mucunoid. In the Mimosaceae, the ability of a new genus (Balizia) to form nodules was reported, as well as nodulation on aerial roots in Inga stipularis. The nodules studied were mainly mucunoid. In the Papilionaceae, nodulation on aerial roots in Poecilanthe hostmannii and on conventional roots of the genus Paramachaerium were reported for the first time. All types of nodular structures were found in this subfamily but the structures were quite uniform at the tribal level. These are consistent with suggestions that nodule morphology has a taxonomic value. Eight functional groups of N2-fixing species are proposed, ranging from light dependance to shade tolerance. These results indicate the important role played by N2-fixing species in the global dynamics of the stand and that N inputs by N2 fixation were continuous along the gradient of energetic resources that characterizes the silvigenetic process. The interactions between the photosynthetic capacities of the species and the ability to fix N2 in low light conditions are discussed.

Leaching of nitrate through soil profiles is a major cause of soil acidification. Legume species differ in their ability to take up nitrate (NO3−) and in the degree to which soil NO3− impairs legume nodulation and N2 fixation. This pot experiment examined species variation in N2 fixation and acid production in grain legumes grown for 6 wk on a sandy soil with Ca(NO3)2 added at 0–56.7 mg N kg−1 soil. The eight legumes studied were Lupinus angustifolius (‘Gungurru’), Lupinus albus (‘Kiev mutant’), Lupinus luteus (‘Teo’), Cicer arietinum (Selection T1587), Pisum sativum (‘Dundale’ and ‘Wirrega’), Vicia faba (‘Fiord’) and Lathyrus sativus (Selection 453). While number of nodules was not significantly affected by treatment with NO3−, increasing supply of NO3− decreased nodule mass, with P. sativum ‘Wirrega’ being most sensitive and C. arientinum and V. faba least sensitive. The concentrations of nitrogen in plants generally decreased as the supply of NO3− increased. Increasing supply of NO3− decreased the percentage of N2 fixation, most significantly in L. sativus and least in L. albus. In the absence of added NO3−, the amounts of H+ produced per unit biomass (specific acid production) differed by as much as 2.2-fold among the species, with C. arientinum generating most H+, followed by L. angustifolius, L. luteus, L. albus, V. faba, L. sativus, P. sativum ‘Dundale’ and P. sativum ‘Wirrega’. This species variation was even greater at higher NO3− supply. In the presence of NO3− at up to 14.2 mg N kg−1, acid production by these legumes correlated well with uptake of excess cations but not with the concentration of nitrogen in plants. For all species, release of H+ declined with increasing supply of NO3−, decreasing most in L. sativus and least in C. arientinum. The results suggest that there might be scope for selecting grain legume species in order to minimize soil acidification by decreasing leaching of NO3−.

Applying silicon in the form of metasilicic acid (H4SiO3) or silicic acid (H4SiO3) to Bradyrhizobium-infected, hydroponically grown cowpea seedlings resulted in a significant (P[les ]0.05) increase in the number of nodules, nodule dry matter, and nitrogen fixed on a per plant basis. Total dry matter of plants increased with silicon supply, and the differences were significant (P[les ]0.05) at the higher silicon concentrations. Cowpea plants cultured in sand were also assessed for their response to silicic acid. Nodule number and nodule mass increased with silicon supply to sand cultured plants, though nitrogen fixation was unaltered. Although silicon is not essential for growth of cowpea, it is important for nodule formation and nodule functioning in hydroponically grown plants. Consequently, data collected and conclusions drawn from earlier glasshouse experiments, which have excluded silicon from nutrient solutions, may be flawed. Future studies on nodulation and nitrogen fixation using legumes in liquid culture must therefore include silicon as a nutrient element.

Seeds of the wetland legume, Lotus uliginosus, were germinated and grown in vermiculite which was either continuously flooded or well-drained. Plants from both treatments were infected by Mesorhizobium loti strain DUS341 via a ‘classical’ root hair pathway, although some flooded plants appeared to be infected via enlarged epidermal cells. Subsequent to infection by M. loti, nodule meristems, which had developed within the root outer cortex, were penetrated by infection threads that released bacteria into the meristematic cells. The infection threads and infection droplets were immunogold labelled with monoclonal antibodies (MAC265 and MAC236) that recognize epitopes (at approx. 155/170 and 170/210 kDa, respectively) on a glycoprotein component of the matrix that surrounded the bacteria within the threads or droplets. Although labelling of infection threads or infection droplets with MAC236 was stronger than that with MAC265, both antibodies strongly labelled material occluding intercellular spaces in the cortices of developing nodules that had not yet expressed nitrogenase (as determined by a lack of signal after immunogold labelling with an antibody raised against nitrogenase component II). After 60 d, nitrogenase activity, shoot and root dry weights, and nodule fresh weight per plant did not differ between the treatments. After a further 30 d submergence, the flooded stems developed extensive aerenchyma and there was profuse development of (nodulated) adventitious roots. Nodules also formed at the junction of adventitious roots and the subtending stem and these were connected vascularly to a small stalk of tissue which gave rise to both a nodule and an adventitious root. The flooded nodules had prominent lenticels, and possible air pathways from the atmosphere to the nitrogen-fixing bacteroids are discussed.

Interactive effects of elevated atmospheric CO2 and arbuscular mycorrhizal (AM) fungi on biomass production and N2 fixation were investigated using black locust (Robinia pseudoacacia). Seedlings were grown in growth chambers maintained at either 350 μmol mol−1 or 710 μmol mol−1 CO2. Seedlings were inoculated with Rhizobium spp. and were grown with or without AM fungi. The 15N isotope dilution method was used to determine N source partitioning between N2 fixation and inorganic fertilizer uptake. Elevated atmospheric CO2 significantly increased the percentage of fine roots that were colonized by AM fungi. Mycorrhizal seedlings grown under elevated CO2 had the greatest overall plant biomass production, nodulation, N and P content, and root N absorption. Additionally, elevated CO2 levels enhanced nodule and root mass production, as well as N2 fixation rates, of non- mycorrhizal seedlings. However, the relative response of biomass production to CO2 enrichment was greater in non-mycorrhizal seedlings than in mycorrhizal seedlings. This study provides strong evidence that arbuscular mycorrhizal fungi play an important role in the extent to which plant nutrition of symbiotic N2-fixing tree species is affected by enriched atmospheric CO2.

The treatment of wheat roots with 2,4-dichlorophenoxyacetic acid (2,4-D) led to the formation of tumour-like structures, para-nodules, which were readily colonized by cyanobacteria. The amount of cyanobacteria found on 2,4-D treated roots was 3.6 times higher than that found on untreated roots. Cyanobacteria penetrated the para- nodules by migrating in between loosely arranged cells that covered their surfaces or by penetrating the space at the junction of root and para-nodule. In plants treated with 2,4-D and co-cultivated with cyanobacteria in medium without combined nitrogen, the rate of acetylene reduction was three times that seen in untreated but colonized roots. Addition of 2,4-D itself did not change the rate of acetylene reduction in free-living culture of Nostoc sp. strain 2S9B. In plants treated with 2,4-D and co-cultivated with Nostoc sp. strain 2S9B in medium without combined nitrogen the nitrogen content of roots but not shoots was significantly increased.

Addition of 2 mM nitrite or ammonium to aerobically incubated cultures of Gloeothece rapidly inhibited N2 fixation (measured as acetylene reduction). In contrast, 2 mM nitrate inhibited N2 fixation less rapidly and less extensively, and often temporarily stimulated nitrogenase activity. The inhibitory effects of both nitrate and ammonium could be prevented by addition of 3 mM L-methionine-DL-sulphoximine, suggesting that the true inhibitor of N2 fixation was an assimilatory product of ammonium rather than either ammonium or nitrate itself. The inhibition of N2 fixation by nitrite could not, however, be prevented by addition of L-methionine-DL- sulphoximine. On the other hand, nitrite (unlike nitrate and ammonium) did not inhibit N2 fixation in cultures incubated under a gas phase lacking oxygen. These findings suggest that the mechanism whereby nitrite inhibits N2 fixation in Gloeothece differs from that of either nitrate or ammonium. The inhibitory effect of nitrite on N2 fixation did not involve reduction of nitrite to nitric oxide, though nitric oxide was a potent inhibitor of nitrogenase activity in Gloeothece. Nitrate and nitrite inhibited the synthesis of nitrogenase in Gloeothece, while ammonium not only inhibited nitrogenase synthesis but also stimulated degradation of the enzyme. In addition, all three compounds favoured the appearance of the Fe-protein of nitrogenase in its larger, presumed inactive, form.

More than 200 angiosperms, distributed in 25 genera, develop root nodule symbioses (actinorhizas) with soil bacteria of the actinomycetous genus Frankia. Although most soils studied contain infective Frankia, cultured strains are available only after isolation from root nodules. Frankia infects roots via root hairs in some hosts or via intercellular penetration in others. The nodule originates in the pericycle. The number of nodules in Alnus is determined by the plant in an autoregulated process that, in turn, is modulated by nutrients such as nitrogen and phosphate. Except in the genera Allocausarina and Casuarina, Frankia in nodules develops so-called vesicles where nitrogenase is localized. Sporulation of Frankia occurs in some symbioses. As a group, actinorhizal plants show a large range of anatomical and biochemical adaptations in order to balance the oxygen tension near nitrogenase. In symbioses with well aerated nodule tissue like Alnus, the vesicles have a multilayered envelope composed mainly of lipids, bacterio-hopanetetrol and their derivatives. This envelope is assumed to retard the diffusion of oxygen into the nitrogenase-containing vesicle. In symbioses like Casuarina, the infected plant cells themselves, rather than Frankia, appear to retard oxygen diffusion, and high concentrations of haemoglobin indicate an infected region with a low oxygen tension. At least in Alnus spp., ammonia resulting from N2 fixation is assimilated by glutamine synthetase in the plant. The carbon compound(s) used by Frankia in nodules is not yet known. Nitrogenase activity decreases in response to a number of environmental factors but recovers upon return to normal conditions. This dynamism in nitrogenase activity is often explained by loss and recovery of active nitrogenase and has been traced to loss and recovery of the nitrogenase proteins themselves. Recovery is partly due to growth of Frankia and to development of new vesicles in the Alnus nodules. In the field, varying conditions continuously affect the plants and the measured rate of N2 fixation is a result not only of the conditions prevailing at the moment but also of the conditions experienced over preceding days. N2 fixed by actinorhizal plants is substantial and actinorhizal plants have great potential in soil reclamation and in various types of forestry. Several species are also useful in horticulture.